All living things use proteins as building blocks in the construction of
their physical forms.In turn,
proteins are composed of folded strands of 20 different smaller subunits called
"amino acids". All amino acids, except for one (glycine), come in two
different forms known as the levoratory (L - left) and dextrorotary (D - right)
forms.These two forms are called
"enantiomers", "chirals", or "stereoisomers",
which basically means that they have the same molecular and structural formula
but cannot be superimposed on each other no matter how they are oriented in
space.In other words, they are
like one's left and right
hands, which are mirror images of each other, but cannot be superimposed onto one
another.

What is especially interesting about these two L- and D-forms, at least for the purposes of this topic, is that the vast
majority of living things only use the L-form.However, as soon as the creature dies, the L-amino acids start to
spontaneously convert to the D-form through a process called
"racemization".If the rate of conversion can be determined, this process of
racemization might be useful as a sort of "clock" to determine the time of death.

In
order to use the rate of racemization as a clock to accurately estimate when a
living thing died, one must know how various
environmental factors may have affected the rate of change from the L- to the
D-form.As it turns out, this rate,
which is different for each type of amino acid, is also exquisitely sensitive to
certain environmental factors.These
include:

Temperature

Amino
acid composition of the protein

Water
concentration in the environment

pH
(acidity/alkalinity) in the environment

Bound
state versus free state

Size
of the macromolecule, if in a bound state

Specific
location in the macromolecule, if in a bound state

Contact
with clay surfaces (catalytic effect)

Presence
of aldehydes, particularly when associated with metal ions

Concentration
of buffer compounds

Ionic
strength of the environment

Of
these, temperature is generally thought to play the most significant role in
determining the rate of racemization since a 1o
increase in temperature results in a 20-25% increase in the racemization rate.1,2
Clearly, this factor alone carries with it a huge potential for error. Even
slight ranges of error in determining the "temperature history" of a
specimen will result in huge "age" calculation errors.

Calibrating
for even a known temperature history also seems to be rather problematic.Consider that the rate of racemization for various amino acids is
determined by placing a protein into a very high temperature environment
(between 95o
and 150o C) and
then extrapolating these results to low temperature environments.1

Such
extrapolations have been fairly recently (1999) called into question by
experiments showing that models based on high temperature kinetics fail to
predict racemization kinetics at physiologic temperatures (i.e., 37o C). The
authors of this particular paper went on to suggest that, "As conformation
strongly influences the rate of Asu [cyclic succinimide] formation and hence Asx
[aspartic acid + asparagine] racemization, the use of extrapolation from high
temperatures to estimate racemization kinetics of Asx in proteins below their
denaturation temperature is called into question . . . We argue that the D:L
ratio of Asx reflects the proportion of non-helical to helical collagen "3Others, such as Collins and Riley have commented in the press that,
"racemization
in free amino acids has, unfortunately, little bearing on observed racemization
of archaeological biominerals."1

Other
experiments have shown that the L- and D-forms of the same amino acid do not
racemize at the same rate.This
means that the equilibrium ratio may be off from "50:50" by as much as
25%.4

As
far as pH is concerned, it seems that the temperature of a solution also affects
the pH of that solution.So, the
amino acid racemization (AAR) rates not only change with the effects of
temperature, but also with the concurrent effects of pH changes, which are
themselves affected by temperature.1 This can only increase the
potential range of error for age determinations.The local buffering effects of bone and shell matrixes are supposed to
limit this effect, but it is still something to consider as potentially
significant when acting over the course of tens of thousands to millions of
years.

Also,
the actual physical structure of an intact protein significantly affects the
rate of racemization of various amino acids.In fact, in many cases this may even be a more significant factor than
the temperature history.As it
turns out, the N-terminal amino acids racemize faster than the C-terminal amino
acids of the same types.Also, the
surface amino acids racemize much faster than the interior amino acids.And, interestingly enough, free amino acids have the slowest racemization
rate of all.Studies with short
peptides have shown that, "replacement
of the asparagine residue with aspartic acid resulted in a 34-fold
decrease in the rate of succinimide (Asu) formation. In the position carboxyl to
asparagine in the peptide the replacement of glycine with a bulky amino acid
such as proline or leucine resulted in a 33-50-fold decrease in the rate
of deamidation"1 [Emphasis added] This clearly emphasizes the
rather dramatic importance that amino acid position and overall protein amino
acid sequence has on the racemization rate.

Hydrolysis,
or the process of breaking down a protein into smaller and smaller fragments,
clearly affects the rate of racemization. The rate itself of hydrolysis
"depends on the strength of the individual peptide bonds, which in turn is
determined by the characteristics of the amino acids on either side of the bond,
the presence of water and the temperature."1 With increased
hydrolysis, the overall rate of the whole specimen would increase since there
are more terminal end and surface amino acids to undergo higher rates of
racemization.All of these are
confounding factors, which, if not known exactly over extended periods of time,
would play havoc with any sort of age determinations. Even the process of
preparing a specimen for racemic dating can affect the D/L ratio.

In
this light, consider that age determinations are usually performed on specimens
for which the amino acid order of the original proteins is unknown.For example, consider that neither the structure nor the proportion of
the amino acids used for dating coral, ostrich eggshell, or snail shells is
known. Again, those like Goodfriend and Hare (1995) have pointed out the
"difficulties in estimating the amount of asparagine in proteins and
reminded researchers of the dangers of extrapolating from the behaviour of pure
Asn in solution at high temperature, to the behaviour of Asn in proteins
associated with biominerals at ambient temperature . . . Using
a simplified model of 'racemization kinetics' of bone collagen over geological
time Collins shows that almost any value of D-Asx can be obtained by varying the
rates of collagen denaturation and
leaching of the denatured product."1

The Interaction of bacteria and fungi with
organic specimens may also be problematic.Such creatures have various enzymes that can digest various types of
proteins, such as collagen, that are commonly used for dating. Various studies,
such as those dealing with 18th and 19th century burials,
showed "unexpectedly high levels of aspartic acid racemization."The authors "suggested that either biological or
chemical degradation of the tooth collagen might have caused these
results." 1 Also, certain types of bacteria and other creatures
actually produce the D-form instead of the L-form of amino acids.So, special care is obviously needed in order to particularly avoid this
sort of contamination.

"Amino acid dating cannot
obtain the age of the material purely from the data itself. The rate of
racemization can not be standardized by itself because it is too changeable.
Thus, because of the rate problem, this dating technique must rely on other
dating techniques to standardize its findings. As a matter of fact, the ages
obtained from racemization dating must rely on other techniques such as Carbon
14, and if the dating of Carbon 14 is not accurate, racemization dating can
never be certain."6

"The
potential variation in the racemization rate has led some paleoanthropologists
to consider this dating technique relative rather than chronometric.
It is, perhaps, best considered to be a calibrated relative dating technique
which puts it somewhere between relative and chronometric methods."7

See
additional references on the relative nature of amino acid racemization dating
in the addendum section ( Link )

Clearly,
all of the above described variables for amino acid racemization rates create great difficulty for
AAR as a
dating technique.In fact, the
difficulties are so great that this technique cannot be and is not used as any
sort of "absolute" dating technique.So, how is it
thought to be at all helpful?Well,
it is thought to be helpful as a "relative" dating technique.

To overcome the various uncertainties
inherent to amino acid dating, the method must be "calibrated" based
on other more reliable techniques such as radiocarbon dating (carbon 14 dating).What happens is that a specimen from a site is chosen as the
"calibration sample" and both a radiocarbon date as well as a D/L
amino acid ratio is determined.These
values are used to solve for a constant or "k" in the formula used to
estimate ages based on the calibration sample.Of course, the "major assumption required with this approach is that the
average temperature experienced by the 'calibration' sample is representative of
the average temperature experienced by other samples from the deposit."1

Much effort has gone into transforming the
data in various ways to achieve linearity between the D/L ratio and the
calibrated age of the specimens in a given location.At first "cubic transformations"' and then later "power
function transformations" were used that seemed to show a "strong
correlation with time, but did not explain the observed kinetics."

What this basically means is that amino acid
dating is not based on any sort of understanding about how racemization takes
place, but is strictly a function of correlation with other dating techniques,
such as the radiocarbon technique. So, if there is any problem with the basis of
the correlation (i.e., radiocarbon dating) then there will also be the same
problem with amino acid dating.

In
this light, it is interesting to consider what happened in 1974 when some of the
major proponents of amino acid dating (Bada et al) decided to analyze the
Paleo-Indian skeletal material from Del Mar, California.Their estimated age of 48,000 years before present (BP)
"stunned" the archaeological community who generally believed these
bones to be less than 10,000 years old. Bada went on to date other skeletal
specimens between the 35,000 and 48,000 year range with one specimen from
Sunnyvale being dated at an astonishing 70,000 years BP. Then, in the 1980s,
something very interesting happened.

"The
Sunnyvale skeleton and the Del Mar tibia were re-dated using uranium
series dating. This resulted in dates of 8,000 to 9,000 years BP for
Sunnyvale and 11,000 to 11,500 for Del Mar.Conventional plus accelerator mass spectrometry (AMS) radiocarbon
dating (Taylor et al. 1983) was carried out on the Sunnyvale skeleton and
results of between 3,600 and 4,850 years BP were obtained. The original
amino acid extractions from the racemization studies of the Paleo-Indian
remains were independently dated by the AMS radiocarbon method at the
Oxford Radiocarbon Accelerator Unit of Oxford University and the NSF
Accelerator Facility for Radioisotope Analysis, University of Arizona.
Bada et al., (1984) published the Oxford results and Taylor et al., (1985)
published a paper combining the results from both laboratories. The Oxford
dates were all between 4,500 and 8,500 years BP and the Arizona dates were
between 3,000 and 6,600 years BP. Bada et al., (1984) stated that the
Oxford AMS results reveal no clear relationship between the radiocarbon
ages of the various skeletons and the extent of the aspartic acid
racemization. They did note that there appeared to be a direct
relationship between the extent of racemization and the level of
preservation of collagen in the bones. Those samples with the most
racemization had the lowest amino acid content and this poor preservation
of protein would contribute to anomalous AAR results.

Later,
based on AMS radiocarbon dates, Bada (1985) calculated a new value for kasp
for the Californian
samples. He used the Laguna skull and the Los Angeles Man skeleton as
'calibration' samples for this. Using the revised value for kasp
he recalculated the
AAR dates of the other Paleo-Indian samples. They all fell within the
Holocene but had much larger error estimates than those of the AMS values.
Although Bada claimed consistency between AAR and AMS dates others
(Pollard and Heron 1996, p. 228) argue that the dates only appear to be
consistent with one another because of the unacceptably large error range
associated with the AAR dates. Pollard and Heron also point out that there
is poor concordance between the conventional and the AMS radiocarbon dates
and there is no concordance between the uranium series dates and any of
the other dates either. At best three of the four methods put the bones in
the Holocene."1

Because of these problems AAR dating of bone
and teeth (teeth in different locations in the same mouth have been shown to
have very different AAR ages) is considered to be an extremely unreliable
practice even by mainstream scientists. That is because the porosity of bones
makes them more "open" to surrounding environmental influences and
leaching.Specimens that are more "closed" to such problems
are thought to include mollusk shells and especially ratite (bird) eggshells
from the emu and ostrich.Of
course, even if these rather thin specimens were actually "closed"
systems (more so than even teeth enamel) they would still be quite subject to
local temperature variations as well as the other above-mentioned potential
problems. For example, even today "very little is known about the protein structure in ratite
eggshell and differences in primary sequence can alter the rate of Asu formation
by two orders of magnitude [100-fold] (Collins, Waite, and van Duin 1999).
Goodfriend and Hare (1995) show that Asx racemization in ostrich eggshell heated
at 80 oC has complex kinetics, similar to that seen in land snails (Goodfriend
1992). The extrapolation of high temperature rates to low temperatures is known
to be problematic (Collins, Waite, and van Duin 1999). A pilot study would be necessary and a reliable relationship between racemate ratio and
time could remain elusive."1

Also, there is a potential problem with
radiocarbon correlations that is quite
interesting.Note what happens to
the correlation constant (k) with assumed age of the specimen in the following
figures.

Interestingly enough, the racemization
constant or "k" values for the amino acid dating of various specimens
decreases dramatically with the assumed age of the specimens (see figures).5
This means that the rate of racemization was thousands of times (up to 2,000
times) different in the past than it is today.Note that these rate differences include shell specimens, which are
supposed to be more reliable than other more "open system" specimens,
such as wood and bone.

Is this a reasonable assumption?Well, this simply must be true if radiocarbon dating is accurate beyond a
few thousand years.But, what if
radiocarbon gets significantly worse as one moves very far back in time beyond
just a few thousand years?In other
words, what would it mean for one to assume that the k-values remained fairly
constant over time as would seem intuitive?Well, with the k-values plotted out horizontally on the graph, the
calculated ages of the specimens would be roughly affected as follows:5

Current
Fossil Age Assignment

40,000

100,000

350,000

1,000,000

Adjusted
Fossil Age Assignment with horizontal k-values

Figure
1

6,000

11,000

18,000

8,000

Figure
2

5,000

14,000

18,000

14,00

0

Clearly
this is a dramatic adjustment that seems to suggest that amino acid racemization
may be more a reflection of the activities of local environmental differences
than any sort of differences in relative ages.This seems especially likely when one considers that each type of
specimen and each different location have different k-values meaning that the
radiocarbon-derived constant in one region or with one type of specimen cannot
be used to calculate the age of any other specimen or even the same type of
specimen in a different location.1

Add
to this the fact that radiocarbon dating is also dependent upon the state of
preservation of the specimen.

"Stafford et al., (1991) discussed AMS radiocarbon dating in
bone at the molecular level. They dated a number of fractions (ranging from
insoluble collagen to individual amino acids) from each of a selection of
differentially preserved mammoth and human bone. Age estimates from the
fractions within a bone were consistent if it was well preserved. They
concluded that a poorly preserved Pleistocene-age fossil >11,000 years in
age would go unrecognised because it would yield a Holocene 14C date. Thus
the final irony is that the poorly preserved Californian Paleo-Indian bones
would return Holocene 14C dates even if they were actually Pleistocene. The
state of preservation of the bone appears to be as important an issue for
radiocarbon dating as it is for AAR dating."1

So,
what do we have? In short, it seems like the claims of some scientists that
amino acid racemization dating has been well established as reliable appears to
be wishful thinking at best. The huge number of confounding factors and a
complete inability to explain the calibrating k-values in terms of amino acid
kinetics leaves those with even a tiny pessimistic bone in their bodies just a
bit underwhelmed.

For
many decades the observation that petroleum shows optical activity, usually
favoring L-enantiomers, was used to prove the biogenic origin of petroleum.
However, more recently there have been scientists who have argued for the non-biogenic
origin of petroleum, citing situations where optical activity can be produced
in non-organic materials, to include hydrocarbons. A particularly
impressive proof of this hypothesis was the discovery of L-enantiomers in
proteins within meteorites ( Link
).

Subsequent
analysis by Bada et. al., challenged this demonstration of
L-enantiomers within meteorites, suggesting that this particular finding is
the likely result of contamination rather than in situ formation
of optical activity within the meteoric proteins ( Link
).

However,
in 1997 research showed that individual amino-acid enantiomers from Murchison
were enriched in the nitrogen isotope 15N relative to their
terrestrial counterparts, which seemed to suggest an extraterrestrial source
for an L-enantiomer excess in the Solar System ( Link
).

Then in 2001 a paper by Pizzarello and Cooper again seemed to confirm the
contamination argument for the origin of optical activity for amino acids
within meteorites. There is currently still some debate, but the
consensus seems to currently favor the contamination theory ( Link
).

This is all very interesting because optical activity decays over time toward
a racemic state. Optically active petroleum is usually found with
temperatures of 66 degrees Celsius. At such temperatures, optical
activity should not be maintained for more than 10 or 20 million years at
most. Yet, optical activity within petroleum, usually of the L-enantiomeric
type, seems to be maintained in significant degrees despite ages assumed to be
over 300 million years old? How is this explained?

Regarding the relative nature of amino acid
racemization dating - i.e., the requirement for calibration against another
dating method for a local area:

"D/L aspartic acid ratios cannot be
converted into an age estimate unless a suitable known age reference
sample is available for calibration of the aspartic acid racemization
rate, e.g., kasp value (Bada 1985; Bada et al. 1979). In the case
of the California paleoindian skeletons, the original racemization ages
were derived using the Laguna skeleton dated by conventional radiocarbon
method (Beta-counting) . . . "

Amino-acid ratios can be used for either
relative or absolute dating. Absolute dating requires calibration with
radiometric techniques, such as radiocarbon dates, and knowledge of the
temperature history of the fossil. Once such information is established
for a region, amino-acid dating may be used with confidence. However,
amino-acid time calibration cannot be extended beyond the area of study
due to regional differences in temperature history.

In those articles I show that after
'calibrating' the amino acid racemization reactions using a radiocarbon
dated bone, it is then possible to date other bones from the same site,
which are either too old or too small for radiocarbon dating. The only
assumption in this approach is that the average temperature experienced
by the calibration sample is representative of the average temperature
experienced by the other sample. Ages thus deduced are in good agreement
with radiocarbon ages determined on the same samples.

To overcome the
problem of inherent uncertainty in the temperature history of sub-fossil
bone Bada and colleagues (Bada and Protsch 1973; Bada et al. 1974)
developed a .calibration" method for dating bones using aspartic
acid racemization. A bone from a site was chosen as a .calibration"
sample and both a radiocarbon date and a D/L aspartic acid ratio were
determined. These values were substituted into equation ( 2.4) on page
15 and it was solved for k. The result was an in situ kasp value
for the site. After substituting in this k asp value equation
(2.4) on page 15 was used to determine the age of other samples from the
site for which only D/L aspartic acid values had been determined. The
major assumption required with this approach is that the average
temperature experienced by the .calibration" sample is
representative of the average temperature experienced by other samples
from the deposit.

More recently researchers have developed
calibration curves using a number of age estimations by independent
dating methods (e. g. radiocarbon) and considerable effort has gone into
transforming data in various ways to achieve linearity in the
relationship between D/L ratio and age. Goodfriend et al., (1992) used a
cubic transformation of D/L data for aspartic acid to achieve linearity.
Later, power function transformations were used for D/L ratios in both
mollusc and ostrich shells (Goodfriend and Hare 1995; Goodfriend 1996).
Such transformations allow a strong correlation with time but do not
explain the observed kinetics.

By determination of the amount of racemization
of aspartic acid in bones from a particular location which have been
dated by the radiocarbon technique, it is possible to calculate the in
situ first-order rate constant for interconversion of the L- and D
enantiomers of aspartic acid. Once this "calibration" has been
calculated, the reaction can be used to date other bones from the
deposit that are either too old to be dated by radiocarbon or that are
too small for radiocarbon dating. The only assumption required with this
approach is that the average temperature experienced by the
"calibration" sample is representative of the average
temperature experienced by older samples.

This study explores time-averaging
(temporal mixing) at very high sampling resolution: that of adjacent
shells collected from the same stratum. Nine samples of the bivalve Chione
fluctifraga were collected from four Holocene cheniers (beach
ridges) on the Colorado Delta (Gulf of California) and 165 shells were
dated using radiocarbon-calibrated amino-acid racemization (D-alloisoleucine/L-isoleucine).

There is a book by a vertebrate
paleontologist on dating methods pertinent to vertebrate paleontologists. It
says little about amino acid racemization, but
simply dismisses it with the comment that it is of little value to
vertebrate paleontology because of its heavy dependence on calibration by
carbon 14.
- Leonard Brand, paleontologist, LLU, personal letter,
December 2007

"Two approaches to calibrating 14C and AAR are used. Interval
calibration involves multiple AAR analyses (>10) of Muliniafrom
previously 14C dated core intervals. Both linear and non-linear
regressions of D/L Asp against 14C age yield comparable R-squared
values (0.91), but the intercept value has not yet been determined by
analysis of modern samples. Direct calibration, currently in progress
for 8 samples, involves both AAR and 14C analysis of separate valves from
articulated Mulinia individuals. Because direct calibration results
are not influenced by time-averaging, they should provide insights into the
reliability of the interval calibration.

Use of AAR data to assess time-averaging not only requires a calibration
curve but also an understanding of all factors that cause a spread in D/L
values for a given core interval. For intervals with >10 analyses, the
coefficient of variation (CV) for D/L Asp is between 3 and 10%, with only
two intervals having CV's >7%. These ranges may be interpreted as
"normal scatter" around an analytical mid-point, with all samples
being essentially the same age, however, time-averaging (as represented by
larger CV's) seems most pronounced within a region of slow or interrupted
deposition at ~1150 cm core depth, between 2800 and 5600 cal yrs BP."
[emphasis added] ( Link)

"Before using AAR ratios in fossils for dating, several tests need to
be completed to ensure that the racemization reaction meets the criteria for
the first order linear reversible or parabolic kenetics needed for dating
(Fig. 4). The two pronged approach using both modern kinetic studies and
calibrated tests on fossils applied to develop AAR dating for ostrich egg
shells is an excellent example.

In high temperature kinetic experiments, the fossil species to be dated is
heated at several different temperatures for various times. After using the
Arrhenius equations (Eq 6) to determine a rate constant for each temperature
tested ( e.g., Fig. 10a), the rate constants are plotted versus temperature
(e.g., Fig. 10b) and extrapolated to Earth surface temperatures. If this
method is used in isolation, then the curve must be assumed to extrapolate
linearly to lower temperatures, an assumption that cannot be guaranteed
without calibration at low temperatures (Blackwell, 1987; Rutter &
Blackwell, 1995).

Calibration tests use several fossils from several diverse sites that have
been accurately and precisely dated by another technique, such as C-14,
etc. . ." [emphasis added]

"With suitable calibration, racemization data can also be used to
construct a history of temperature for the samples in question.
Understanding diagenetic amino acid racemization (AAR) kinetics
requires a combination of laboratory kinetic experimentation and testing of
kinetic models with natural field samples that have independent
chronostratigraphic control. These models must also incorporate information
on the temperature dependence ("apparent" or "bulk"
activation energy) as derived from either elevated temperature laboratory
experiments or data from field samples with known ages and
diagenetic temperatures. Over the past two decades, several well-calibrated
datasets and a variety of kinetic models have become available: foraminifera
results provide calibration for low temperature kinetics ( e.g., Muller,
1984) and paired radiocarbon-AAR results provide calibration at higher
surface temperatures (e.g., Miller et al., 1997)." [emphasis added]

"AAR
is not a numerical dating method, per se; however, it can be used for a
variety of chronological and palaeotemperature applications. Provided there
is some independent age control, AAR can be used to extend or to improve
upon the chronology, or to reconstruct the temperature history at a
site."

"The increase in the proportion of D-amino acids (a function of time
and temperature) can be used as a tool for estimating age. Attempts to
provide absolute dates calibrate the amount of racemization using samples
of known age and then use these predict the age of samples with known
D/L values but of unknown age. A recent paper by Kaufman (in press Geology)
estimates that the compound accuracy of this approach is +/- 20%."
[accessed 11/29/07] ( Link)

"Although the finding of this dating method was meaningful, this
dating method is considered controversial. The rate of racemization is
dependent on temperature, or the thermal history of the fossil. The rate of
racemization increases at warmer sites than at cooler sites. Samples from
areas of different latitude tend to have greater age range differences. To
obtain accurate dates, the temperature has to be constant for thousands
of years. The speed of racemization slows down if the sample materials
gets cold. The study carried out with bone revealed that unassured
temperature of plus or minus 2 degree will cause an plus or minus 50% error
of the age.

Amino
acid dating cannot obtain the age of the material purely from the data
itself. The rate of racemization can not be standardized by itself
because it is too changeable. Thus, because of the rate problem, this dating
technique must rely on other dating techniques to standardize its findings.
As a matter of fact, the ages obtained from racemization dating must rely on
other techniques such as Carbon 14, and if the dating of Carbon 14 is not
accurate, racemization dating can never be certain.

Materials can easily be contaminated. Sample material can lose or gain amino
acids by leaching, diagenetic formation of amino acids, bacterial
contamination, and/or contaminated during collection or preparation. If the
materials are contaminated by water, their racemic clocks will be ruined.
Even though there must be some moisture for the occurrence of racemization,
continual inflow of moisture can cause many kinds of contamination. For
example, if pH of the moisture is higher, the racemization process would be
very rapid, and that changes the age of the sample." [Bibliography with
almost all references from 2000] ( Link)

"The potential variation in the racemization rate has led some
paleoanthropologists to consider this dating technique relativerather
than chronometric. It is, perhaps, best considered to be a calibrated
relative dating technique which puts it somewhere between
relative and chronometric methods." [accessed 11/29/07] ( Link)

"If the racemization rate for a particular system in any given material
is known then AAR can be used to estimate the thermal history of any
cross-dated samples. However, good independent age controls such as
14C dated samples and a reliable kinetic model for racemization rates are
needed (Johnson and Miller 1997)."